Mentor Tasks for the new push on protein expression using Sadhana's MtPSTP and Joey and Chrsitina's FtHAP
the students will each do a 500ml expression on their own - but we need to make some reagents for them
-- they can see the Targets page for which enzyme they need to do
I have ordered TCEP, PMSF, B-Per, Imidazole, etc. - when these arrive, label them and store appropriately (should say on bottle how to stores, otherwise look up on web)
Can see the Solutions & Dilutions spreadsheet in GDocs/Misc/ for some help with calculations and Molecular Weights
SOLUTIONS TO MAKE: LARRY Make LB - 2Liters Make 5 M NaCl - 500 ml in water, sterile filter into a bottle, label, store at Room Temp
SADHANA Make Kanamycin- stocks 50ug/ul store in -20degC, Make 30 tubes of 500 ul each. Sterile filter with syringe filterbefore aliquoting into tubes. Label. Store in -20degC
Make IPTG - 1 M IPTG stock to make. Make 30 tubes of this with 500 ul in each. Label. Store in -20degC
CHRISTINA Make Lysozyme - 20 aliquots of 500 ul at 50 mg/ml in 1.7 ml tubes. Label. Store in -20degC
Make TCEP - make ~50 x 100ul aliquots of 500 mM aliquots medium size centrifuge tubes. Label, put in a box and Store at Room Temp in cubby.
JOEY Make PMSF - 30 x 500 ul aliquots of 10mM in Isopropanol 1.7 ml tubes. Label. store in -20degC
Make Imidazole - 1M stock make in beaker of 500 ml of water, then bottle top sterile filter it into a bottle, store at Room Temp
JC
When you work on Friday - work on whatever from above does not get done on Thursday and the Lysis Buffer:
NEW Lysis Buffer for Sonication (make 500 ml) (see GDocs 'Solutions&Dilutions' in /Misc folder for calculations) Tris 100 mM NaCl 300 mM Imidazole 30 mM pH 8.0 w/ NaOH label, store at 4degC
Later: Make Compounds in DMSO - 50 uM in 100% DMSO, then 5 or 10 uM
NEW Wash and Elution Buffers and Storage Buffer(see GDocs 'Solutions&Dilutions' in /Misc folder for calculations)
Wash Buffer Tris 100 mM NaCl 300 mM Imidazole 30 mM pH 8.0 w/ NaOH
Elution Buffer Tris 100 mM NaCl 300 mM Imidazole 300 mM pH 8.0 w/ NaOH
FPLC column buffer - tris Also 'storage' buffer**
Tris 50 mM
NaCl 150 mM
pH 8.0 w/ NaOH
Then filter sterilize
Then degass
make glycerol stocks of E. coli competent cells (DH5alpha and BL21(DE3) - make plates with no resistance, grow up 4ml culture, see Glycerol Stocks protocol for freezing down.
-- there is also a protocol using DMSO in the Molecular Cloning books
Midiprep pGBR22 (for the students first RE digest protocol)
Teaching Points for Labs Spring 12
Larry - create Wikispaces pages for each student for Protein Labs - Done - Thanks!
J.C. - '20 questions' form - Done - Thanks!
Christina - script for Vina, script to put compound ID label in right place on SDF - Done - Thanks!
Michael - pseudoknot stuff, Mini Research Wiki post templates Done - Thanks!
022812
Laptop setup - Michael & Christina - Great job on this guys. A huge help!
Put a tape label on cover of each laptop with name and FRI on it and room # (PAI 2.14).
Put a tape label on each mouse and charger (these don't need to be tied to an individual laptop - but just say "FRI PAI 2.14" on them Computer names:
Valine- Dr. B put on ViiA7 and did some Windows updates
Aspartate
Serine
Tryptophan
Histidine
Methionine Computer Description: FRI Laptop Username: VDSclass
no password until you are done installing everything
Set up UT Restricted Wireless - sign into GUEST first then open a webpage to get forced to the Restricted installer
Firefox
Internet Explorer - Boo!
Chrome
- Bookmarks to install on browsers:
Gmail, UTexas, CNS, FRI, VDS Google Site, VDS Wikispaces, RT-qPCR Course, Residential Halls Study Group, PDB, BRENDA, MolPROBITY
Add password:
same as our Google Docs login
Low priority: put an FRI logo as the Desktop. Maybe try this one:
02/26/12: EXPRESSION LAB:
Also, I believe we will be starting the Protein Expression lab early at the end of this week (Thurs & Fri). Basically we will have a few students go this week and just do the Transformation. If you remember - it is a 3 day experiment. So, it is important to spread them out early since we have 35 students!
That 1st step is only a transformation - so, you guys have done like a million of those!
SCHEDULING EXPRESSION LAB:
Also, I am going to be out of town BOTH this Friday and Next.
So, for this week on:
Thursday - Joey will be in charge of handling the Protein Expression Lab
Friday - Joshua will be in charge of handling the Protein Expression Lab
Adam - we might need you to hang around longer on Friday if you can.
Let me know if this does not work and we need to make schedule switches.
I will post the lab for the Protein Expression Lab late this week.
MATERIALS FOR EXPRESSION LAB:
Sadhana - I think that you remade the LB -so we should be good to go with that for now.
Larry - I would like to have you check on whether we have enough plates for that lab as an extra task for you this week.
--- First check and see what we have. Then make what we are short on.
In total we need 35*2 = 70 Ampicillin plates and 35 No Antibiotic plates. - Done by Larry - Thanks!
Ideally you could do this on Monday or Tuesday. At the minimum, we will need several of each by Thursday.
Thanks,
Dr. B
012512- Adam to buy at BioSci storeroom
EDTA (sigma) Cat # E4884
3 medium lab coats (cloth)
squirt bottles - 2 of these (for DI water and nanopure water)
spray bottles - 2 of these
Sharpie pens (black, fine tip) - 6 of these
labeling tape (thick and thin) - various colors - maybe about 10 rolls
Larry - did you do a gel of PTP1b or of YopH the other day? - Thanks, Dr. B
Wed Jan 18th
Adam - grow up YopH pNIC-Bsa4 DH5alpha colony in 2x80ml of LB overnight (use water bath shakers since we don't trust the air shaker)
Joshua - grow up 2x 25 ml of pGBR22 in BL21(DE3) - use Michael's plate
ToDo List (overall)
Clean out Fridges (silver and white) -- discard old/moldy plates, old tubes Move all Staff plates to the White fridge\
move powdered chemicals in large buckets to bottles generate a mentors hours log sheet generate a time sheets due date sheet (from UT webpage) make name badges and print make -80degC freezer signs (laminate them with new laminator!)
make lysozyme - store in freezer
make 1M IPTG - store in freezer
make Kanamycin stocks 50ug/ul store in -20degC
buy stuff at BioScience storeroom (have to wait til Tuesday to do)
bleach
alconox??
pasteur pippettes
rubber bubls for pasteur pipettes
Monday Jan 16th
Team Building Activity Staff picture Buffer Titration lab (if we have pH probes from other FRI stream) Beer's Law Lab
PyMol Labs
Purify/Characterize pGBR22 o/n cultures Generate time sheets
Calibrate pH meter: ALL clean glassware: ALL clean out old plates and transfer important ones to white fridge: ALL
Sadhana - Midiprep PSTP pNIC-Bsa4 in DH5alpha
Larry - Midiprep YopH pNIC-Bsa4 in DH5alpha
Autoclave colirollers
divide up colirrolers in to 6 each in sterile tubes of (600 ul tubes) -Done by Joshua
Make LB/Agar plates with NO antibiotic (for sneeze plates) - need about 35 of these
Post RT-qPCR signs in NHB and MBB
Wolbachia project
Biooscientific project
Sunday Jan 15th
Larry add full labels to Midiprep DNA tubes
Sadhana - check protein gel
Beer's Law: Christina + Joshua
Buffer Titration: none - no probes
Transformation of pGBR22 in BL21(DE3) : Michael
Grow up pGBR22 in BL21(DE3) overnight - ALL
Sadhana - Friday Jan 13th
YopH in BL21(DE3), purify, characterize
PTP1b in BL21(DE3), purify, characterize
Midiprep - PSTP
Larry (Wed/Thursday)
Express YopH and PTP1B in 1 L of LB each - DONE
-- grow to .1 OD then to 0.3 OD (add IPTG) - wait 4 hrs, spin down and store in -80degC
Spin down Midiprep DNA colonies that Sadhana grew up (maybe Midiprep them) - DONE
Midiprep (YopH-pNicBsa4, pBGR22) - DONE
Troubleshoot Labs
-Wet Labs (Pipetting step of Buffers & Solutions, Beer's Law, Buffer Titration)
-
[[image:/i/file_not_found.png width="32" height="32" caption="File Not Found"]]File Not Found
-Gold labs (if they have been revised by then)
TUES night: Dr. B -
Grow up 2x 80 ml of LB o/n with Kan - for Midiprep (Sadhana- did we use DH5alpha or BL21's for this? - Thanks, Dr. B 1/10/12)
-YopH in pNIC-Bsa4 - J.C. - we have a plate of this as DH5alpha colonies
-PSTP in pNIC-Bsa4 - Sadhana - we have a plate of this as DH5alpha colonies
-grow up pGBR22 in DH5alpha o/n
Express small culture o/n of YopH and PTP1b
Sadhana - Monday Jan 9th
Make LB media
Make some Kan plates, Kan Suc, and Amp
Transformations of plasmids in DH5alpha -pGBR22 in pGEM-T - have DNA (use AMP plates)
Transformations of plasmids in (BL21(DE3)) -YopH in pNIC-Bsa4 - J.C.- we have Mini-prep DNA in Final Plasmids box -PSTP in pNIC-Bsa4 - Sadhana - we have Mini-prep DNA in Final Plasmids box -pGBR22 in pGEM-T - have DNA (use AMP plates) -PTP1b in pNIC-Bsa4
Troubleshoot Labs
-Wet Labs (Pipetting step of Buffers & Solutions)
-PyMol (1 & 2 & 3) -
[[image:/i/file_not_found.png width="32" height="32" caption="File Not Found"]]File Not Found
&
[[image:/i/file_not_found.png width="32" height="32" caption="File Not Found"]]File Not Found
Storage for Spin Tube Concentators: the green arrow denotes where the Ethanol should go up to. The red arros is too low - it will dry out the membrane.
Plasmids that we have as Midi Prep
SfApy in pUC19 - Indra
SalCA in pNIC-Bsa4 - Krishna
MPTPB in pNIC-Bsa4 - Joshua
FtHap in pNIC-Bsa4 - Christina
Plasmids that we need to transform (from Mini-prep DNA)
YopH in pNIC-Bsa4 - J.C.- we have Mini-prep DNA in Final Plasmids box
PSTP in pNIC-Bsa4 - Sadhana - we have Mini-prep DNA in Final Plasmids box
PP2C in pNIC-Bsa4 - Justin
DH5alpha colonies that we need to grow up 2x 80 ml of LB o/n with Kan - for Midiprep
YopH in pNIC-Bsa4 - J.C. - we have a plate of this
PSTP in pNIC-Bsa4 - Sadhana - we have a plate of this
Colirollers - autoclave and distribute to small 600 ul tubes (~6 per tube)
RADHIKA: LB/Kan/Suc plates
REAGENTS THAT WE NEED: For new Lysis Buffer 50 mg/ml Lysozyme in 500 ul aliquots in 1.7 ml tubes. Store in -20degC
ZOE: TCEP aliquots (already at 500 mM - just move from glass vials to tubes and store at Room Temp in cubby)
YILING: PMSF in 500 ul aliquots in 1.7 ml tubes (10mM concentration in isopropanol) store in -20degC
YILING: 1M Imidazole (make in water) store at Room Temp
ZOE: 5 M NaCl (make in water) store at Room Temp
NEW Wash and Elution Buffers and Storage Buffer(see GDocs 'Solutions&Dilutions' in /Misc folder for calculations)
Wash Buffer Tris 100 mM NaCl 300 mM Imidazole 30 mM pH 8.0 w/ NaOH
Elution Buffer Tris 100 mM NaCl 300 mM Imidazole 300 mM pH 8.0 w/ NaOH
FPLC column buffer - tris Also 'storage' buffer**
Tris 50 mM
NaCl 150 mM
pH 8.0 w/ NaOH
Then filter sterilize
Then degass
For Fall 12 Semester:
103112
VDS Mentors are the best!
Mentor Tasks for the new push on protein expression using Sadhana's MtPSTP and Joey and Chrsitina's FtHAP
the students will each do a 500ml expression on their own - but we need to make some reagents for them
-- they can see the Targets page for which enzyme they need to do
I have ordered TCEP, PMSF, B-Per, Imidazole, etc. - when these arrive, label them and store appropriately (should say on bottle how to stores, otherwise look up on web)
Can see the Solutions & Dilutions spreadsheet in GDocs/Misc/ for some help with calculations and Molecular Weights
SOLUTIONS TO MAKE:
LARRY
Make LB - 2Liters
Make 5 M NaCl - 500 ml in water, sterile filter into a bottle, label, store at Room Temp
SADHANA
Make Kanamycin- stocks 50ug/ul store in -20degC, Make 30 tubes of 500 ul each. Sterile filter with syringe filterbefore aliquoting into tubes. Label. Store in -20degC
Make IPTG - 1 M IPTG stock to make. Make 30 tubes of this with 500 ul in each. Label. Store in -20degC
CHRISTINA
Make Lysozyme - 20 aliquots of 500 ul at 50 mg/ml in 1.7 ml tubes. Label. Store in -20degC
Make TCEP - make ~50 x 100ul aliquots of 500 mM aliquots medium size centrifuge tubes. Label, put in a box and Store at Room Temp in cubby.
JOEY
Make PMSF - 30 x 500 ul aliquots of 10mM in Isopropanol 1.7 ml tubes. Label. store in -20degC
Make Imidazole - 1M stock make in beaker of 500 ml of water, then bottle top sterile filter it into a bottle, store at Room Temp
JC
When you work on Friday - work on whatever from above does not get done on Thursday and the Lysis Buffer:
NEW Lysis Buffer for Sonication (make 500 ml)
(see GDocs 'Solutions&Dilutions' in /Misc folder for calculations)
Tris 100 mM
NaCl 300 mM
Imidazole 30 mM
pH 8.0 w/ NaOH
label, store at 4degC
Later:
Make Compounds in DMSO - 50 uM in 100% DMSO, then 5 or 10 uM
NEW Wash and Elution Buffers and Storage Buffer(see GDocs 'Solutions&Dilutions' in /Misc folder for calculations)
Wash Buffer
Tris 100 mM
NaCl 300 mM
Imidazole 30 mM
pH 8.0 w/ NaOH
Elution Buffer
Tris 100 mM
NaCl 300 mM
Imidazole 300 mM
pH 8.0 w/ NaOH
FPLC column buffer - tris Also 'storage' buffer**
Tris 50 mM
NaCl 150 mM
pH 8.0 w/ NaOH
Then filter sterilize
Then degass
make glycerol stocks of E. coli competent cells (DH5alpha and BL21(DE3) - make plates with no resistance, grow up 4ml culture, see Glycerol Stocks protocol for freezing down.
-- there is also a protocol using DMSO in the Molecular Cloning books
Midiprep pGBR22 (for the students first RE digest protocol)
FROM SPRING 2012:
Please Enter your class schedules on the Google Docs Spreadsheet (SchedulingMentorsforGDOcsSp12)
Teaching Points for Labs Spring 12
Larry - create Wikispaces pages for each student for Protein Labs - Done - Thanks!
J.C. - '20 questions' form - Done - Thanks!
Christina - script for Vina, script to put compound ID label in right place on SDF - Done - Thanks!
Michael - pseudoknot stuff, Mini Research Wiki post templates Done - Thanks!
022812
Laptop setup - Michael & Christina - Great job on this guys. A huge help!
Put a tape label on cover of each laptop with name and FRI on it and room # (PAI 2.14).Put a tape label on each mouse and charger (these don't need to be tied to an individual laptop - but just say "FRI PAI 2.14" on them
Computer names:
Valine- Dr. B put on ViiA7 and did some Windows updates
Aspartate
Serine
Tryptophan
Histidine
Methionine
Computer Description: FRI Laptop
Username: VDSclass
no password until you are done installing everything
- Set up UT Restricted Wireless - sign into GUEST first then open a webpage to get forced to the Restricted installer
*Download 'RemoveCredentials.exe' to DesktopSoftware to be installed:
- Microsoft Forefront Client Security - Bevoware
Secure Shell - BevowarePutty
Xming - with MESA and Xming Fonts:
http://www.straightrunning.com/XmingNotes/
- Microsoft Office 2007 - Dr. B has CD
PyMol - on our Google DocsWinSCP
7-zip
Logger Pro ve. 8.3 - Dr. B has USB thumb drive with this
- Viia7 v 1.2 software for RT-qPCR - install software - then get key from Dr. B
http://www.appliedbiosystems.com/absite/us/en/home/support/software/real-time-pcr/viia7-software.htmlFirefox
Internet Explorer - Boo!
Chrome
- Bookmarks to install on browsers:
Gmail, UTexas, CNS, FRI, VDS Google Site, VDS Wikispaces, RT-qPCR Course, Residential Halls Study Group, PDB, BRENDA, MolPROBITY
Add password:
same as our Google Docs login
Low priority: put an FRI logo as the Desktop. Maybe try this one:
02/26/12:
EXPRESSION LAB:
Also, I believe we will be starting the Protein Expression lab early at the end of this week (Thurs & Fri). Basically we will have a few students go this week and just do the Transformation. If you remember - it is a 3 day experiment. So, it is important to spread them out early since we have 35 students!
That 1st step is only a transformation - so, you guys have done like a million of those!
SCHEDULING EXPRESSION LAB:
Also, I am going to be out of town BOTH this Friday and Next.
So, for this week on:
Thursday - Joey will be in charge of handling the Protein Expression Lab
Friday - Joshua will be in charge of handling the Protein Expression Lab
Adam - we might need you to hang around longer on Friday if you can.
Let me know if this does not work and we need to make schedule switches.
I will post the lab for the Protein Expression Lab late this week.
MATERIALS FOR EXPRESSION LAB:
Sadhana - I think that you remade the LB -so we should be good to go with that for now.
Larry - I would like to have you check on whether we have enough plates for that lab as an extra task for you this week.
--- First check and see what we have. Then make what we are short on.
In total we need 35*2 = 70 Ampicillin plates and 35 No Antibiotic plates. - Done by Larry - Thanks!
Ideally you could do this on Monday or Tuesday. At the minimum, we will need several of each by Thursday.
Thanks,
Dr. B
012512- Adam to buy at BioSci storeroom
EDTA (sigma) Cat # E48843 medium lab coats (cloth)
squirt bottles - 2 of these (for DI water and nanopure water)
spray bottles - 2 of these
Sharpie pens (black, fine tip) - 6 of these
labeling tape (thick and thin) - various colors - maybe about 10 rolls
Larry - did you do a gel of PTP1b or of YopH the other day? - Thanks, Dr. B
Wed Jan 18th
Adam - grow up YopH pNIC-Bsa4 DH5alpha colony in 2x80ml of LB overnight (use water bath shakers since we don't trust the air shaker)Joshua - grow up 2x 25 ml of pGBR22 in BL21(DE3) - use Michael's plate
ToDo List (overall)
Clean out Fridges (silver and white)
-- discard old/moldy plates, old tubes
Move all Staff plates to the White fridge\
move powdered chemicals in large buckets to bottles
generate a mentors hours log sheet
generate a time sheets due date sheet (from UT webpage)
make name badges and print
make -80degC freezer signs (laminate them with new laminator!)
make lysozyme - store in freezer
make 1M IPTG - store in freezer
make Kanamycin stocks 50ug/ul store in -20degC
buy stuff at BioScience storeroom (have to wait til Tuesday to do)
bleach
alconox??
pasteur pippettes
rubber bubls for pasteur pipettes
Monday Jan 16th
Team Building ActivityStaff picture
Buffer Titration lab (if we have pH probes from other FRI stream)
Beer's Law Lab
PyMol Labs
Purify/Characterize pGBR22 o/n cultures
Generate time sheets
Calibrate pH meter: ALL
clean glassware: ALL
clean out old plates and transfer important ones to white fridge: ALL
Sadhana - Midiprep PSTP pNIC-Bsa4 in DH5alpha
Larry - Midiprep YopH pNIC-Bsa4 in DH5alpha
Autoclave colirollers
- divide up colirrolers in to 6 each in sterile tubes of (600 ul tubes) -Done by Joshua
Make LB/Agar plates with NO antibiotic (for sneeze plates) - need about 35 of thesePost RT-qPCR signs in NHB and MBB
Wolbachia project
Biooscientific project
Sunday Jan 15th
- Larry add full labels to Midiprep DNA tubes
- Sadhana - check protein gel
Beer's Law: Christina + JoshuaBuffer Titration: none - no probes
Transformation of pGBR22 in BL21(DE3) : Michael
Grow up pGBR22 in BL21(DE3) overnight - ALL
Sadhana - Friday Jan 13th
Larry (Wed/Thursday)
Express YopH and PTP1B in 1 L of LB each - DONE-- grow to .1 OD then to 0.3 OD (add IPTG) - wait 4 hrs, spin down and store in -80degC
Spin down Midiprep DNA colonies that Sadhana grew up (maybe Midiprep them) - DONE
Midiprep (YopH-pNicBsa4, pBGR22) - DONE
Troubleshoot Labs
-Wet Labs (Pipetting step of Buffers & Solutions, Beer's Law, Buffer Titration)
-
[[image:/i/file_not_found.png width="32" height="32" caption="File Not Found"]]File Not Found
-Gold labs (if they have been revised by then)
TUES night: Dr. B -
Grow up 2x 80 ml of LB o/n with Kan - for Midiprep (Sadhana- did we use DH5alpha or BL21's for this? - Thanks, Dr. B 1/10/12)-YopH in pNIC-Bsa4 - J.C. - we have a plate of this as DH5alpha colonies
-PSTP in pNIC-Bsa4 - Sadhana - we have a plate of this as DH5alpha colonies
-grow up pGBR22 in DH5alpha o/n
Express small culture o/n of YopH and PTP1b
Sadhana - Monday Jan 9th
Make LB mediaMake some Kan plates, Kan Suc, and Amp
Transformations of plasmids in DH5alpha
-pGBR22 in pGEM-T - have DNA (use AMP plates)
Transformations of plasmids in (BL21(DE3))
-YopH in pNIC-Bsa4 - J.C.- we have Mini-prep DNA in Final Plasmids box
-PSTP in pNIC-Bsa4 - Sadhana - we have Mini-prep DNA in Final Plasmids box
-pGBR22 in pGEM-T - have DNA (use AMP plates)
-PTP1b in pNIC-Bsa4
Troubleshoot Labs
-Wet Labs (Pipetting step of Buffers & Solutions)
-PyMol (1 & 2 & 3) -
[[image:/i/file_not_found.png width="32" height="32" caption="File Not Found"]]File Not Found
&
[[image:/i/file_not_found.png width="32" height="32" caption="File Not Found"]]File Not Found
Mentor Contract on GDocs
Timesheet Pay Periods and Due Dates (see bottom of link)
OLD STUFF (Fall 2011):
Tris-acetate buffer that is 500 mM pH 5.5 - see Solutions&DilutionsforGdocs spreadsheetLB media - autoclaved
Make ProtocolHomemadeSDSPageGelSolutionsVDS_Fall11.docx gels
Plasmids that we have as Midi Prep
SfApy in pUC19 - Indra
SalCA in pNIC-Bsa4 - Krishna
MPTPB in pNIC-Bsa4 - Joshua
FtHap in pNIC-Bsa4 - Christina
Plasmids that we need to transform (from Mini-prep DNA)
YopH in pNIC-Bsa4 - J.C.- we have Mini-prep DNA in Final Plasmids box
PSTP in pNIC-Bsa4 - Sadhana - we have Mini-prep DNA in Final Plasmids box
PP2C in pNIC-Bsa4 - Justin
DH5alpha colonies that we need to grow up 2x 80 ml of LB o/n with Kan - for Midiprep
YopH in pNIC-Bsa4 - J.C. - we have a plate of this
PSTP in pNIC-Bsa4 - Sadhana - we have a plate of this
Colirollers - autoclave and distribute to small 600 ul tubes (~6 per tube)
RADHIKA: LB/Kan/Suc plates
REAGENTS THAT WE NEED:
For new Lysis Buffer
50 mg/ml Lysozyme in 500 ul aliquots in 1.7 ml tubes. Store in -20degC
ZOE: TCEP aliquots (already at 500 mM - just move from glass vials to tubes and store at Room Temp in cubby)
YILING: PMSF in 500 ul aliquots in 1.7 ml tubes (10mM concentration in isopropanol) store in -20degC
YILING: 1M Imidazole (make in water) store at Room Temp
ZOE: 5 M NaCl (make in water) store at Room Temp
NEW Wash and Elution Buffers and Storage Buffer(see GDocs 'Solutions&Dilutions' in /Misc folder for calculations)
Wash Buffer
Tris 100 mM
NaCl 300 mM
Imidazole 30 mM
pH 8.0 w/ NaOH
Elution Buffer
Tris 100 mM
NaCl 300 mM
Imidazole 300 mM
pH 8.0 w/ NaOH
FPLC column buffer - tris Also 'storage' buffer**
Tris 50 mM
NaCl 150 mM
pH 8.0 w/ NaOH
Then filter sterilize
Then degass